Magnolol inhibits migration of vascular smooth muscle cells via cytoskeletal remodeling pathway to attenuate neointima formation

BackgroundIncreased proliferation and migration of vascular smooth muscle cells (VSMCs) contribute importantly to the formation of both atherosclerotic and restenotic lesions. The objective of this study was to investigate the effect of magnolol on VSMC migration.MethodsThe proteolytic activity of matrix metalloproteinases (MMPs) in tumor necrosis factor alpha (TNF-α) stimulated VSMCs was performed by gelatin zymography. VSMC migration was assessed by wound healing and Boyden chamber methods. Collagen induced VSMC adhesion was determined by spectrofluorimeter and stress fibers formation was evaluated by fluorescence microscope. The expression of signaling molecules involved in stress fibers formation was determined by western blot. The phosphorylation of myosin light chain (MLC20) was determined by urea-glycerol polyacrylamide gel electrophoresis. Immunohistochemistry was performed to determine the expression of β1-integrin and collagen type I in the injured carotid arteries of rats on day 35 after vascular injury.ResultsVSMC migration was strongly inhibited by magnolol without affecting MMPs expression. Also, magnolol inhibited β1-integrin expression, FAK phosphorylation and RhoA and Cdc42 activation to inhibit the collagen induced stress fibers formation. Moreover, magnolol inhibited the phosphorylation of MLC20. Our in vivo results showed that magnolol inhibited β1-integrin expression, collagen type I deposition and FAK phosphorylation in injured carotid arteries without affecting MMP-2 activity.ConclusionsMagnolol inhibited VSMC migration via inhibition of cytoskeletal remodeling pathway to attenuate neointima formation.General significanceThis study provides a rationale for further evaluation of magnolol for the management of atherosclerosis and restenosis.     

Contrasting responses of migration strategies in two European thrushes to climate change

Migration is a widespread strategy that enables animals to escape harsh winter conditions. It has been well documented that migration phenology in birds is changing in response to recent climate warming in the northern hemisphere. Despite the existence of large temporal and geographical scale ringing data on birds in Europe, changes in migration strategies in relation to climate warming have not been well studied, mainly because of a lack of appropriate statistical methods. In this paper, we develop a method that enables us to investigate temporal changes in migration strategies from recoveries of dead ringed birds. We estimated migration probability as the ratio between recovery probabilities of conspecific birds originating from different countries but potentially wintering in the same country. We applied this method to two European thrushes: the entirely migrant redwing Turdus iliacus , and the partially migrant blackbird T. merula . We tested for an immediate and a 1-year lagged relationship between our migration probability and climatic covariates (i.e. mean winter temperature in France and the North Atlantic Oscillation). Using ringing-recovery data collected in Finland, Germany, Switzerland and France from 1970 to 1999, we detected contrasting responses in these two species, likely related to their different migratory behaviours. Both species showed a decline in the probability for northern and eastern birds to winter in France. The entirely migratory redwing exhibited a year-to-year plastic response to climate, whereas the decline in the partially migrant blackbird was smooth, suggesting underlying genetic processes. The proposed method, thus, allows us to identify useful indicators of climatic impacts on migration strategies, as well as highlighting differences between closely related species.     

c-Cbl regulates migration of v-Abl-transformed NIH 3T3 fibroblasts via Rac1

Cellular events like cell adhesion and migration involve complex rearrangements of the actin cytoskeleton. We have previously shown that the multidomain adaptor protein c-Cbl facilitates actin cytoskeletal reorganizations that result in the adhesion of v-Abl-transformed NIH 3T3 fibroblasts. In this report, we demonstrate that c-Cbl also enhances migration of v-Abl-transformed NIH 3T3 fibroblasts. This effect of c-Cbl depends on its tyrosine phosphorylation, specifically on phosphorylation of its Tyr-731, which is required for binding of PI-3' kinase to c-Cbl. Furthermore, we demonstrate that the effect of c-Cbl on migration of v-Abl-transformed fibroblasts is mediated by active PI-3' kinase and the small GTPase Rac1. Our results also indicate that ubiquitin ligase activity of c-Cbl is required, while spatial localization of c-Cbl to the pseudopodia is not required for the observed effects of c-Cbl on cell migration.     

Monovalent copper as a potential catalyst for formation of acetaldehyde via the migration of methyl radicals to the coordinated carbonyl in the complex (CO)Cu-CH3

Cu_aq⁺ forms stable complexes with carbon monoxide in aqueous solutions. Furthermore it reacts very fast with aliphatic radicals. The reaction of Cu(CO)_maq⁺ with methyl radicals, ^•CH₃ was studied using the pulse-radiolysis technique. The results point out that methyl radicals react with Cu(CO)_aq⁺ to form an unstable intermediate with a Cu^II-C σ bond identified as (CO)Cu^II-CH₃⁺, k = (1.1±0.2) × 10⁹ M⁻¹ s⁻¹. This intermediate has a strong LMCT charge transfer band (λ_max = 385 nm, _max = 2500 M⁻¹ cm⁻¹) which is similar to the absorption bands of other transient complexes with Cu^II-alkyl σ bonds. The coordinated carbon monoxide in (CO)Cu^II-CH₃⁺ inserts into the copper—carbon bond (or rather the coordinated methyl migrates to the coordinated carbon monoxide ligand) at a rate of (3.0±0.8) × 10² s⁻¹ to form the copperacetyl complex (CO)_mCu^II-C(CH₃)=O⁺ (λ_max = 480 nm, _max = 2100 M⁻¹ cm⁻¹). The rate of formation of (CO)Cu^II-CH₃⁺ and of the insertion reaction are pH independent. The complex (CO)_mCu^II-C(CH₃)=O⁺ is also unstable and decomposes heterolytically to yield acetaldehyde and Cu_aq²⁺ as the final stable products. This reaction is slightly pH dependent. The same reactivity pattern has been observed for the Cu(CO_naq⁺ complexes (n = 2 or 3). The results clearly point out that CO remains coordinated to transient complexes of the type Cu^II-alkyl.     

The effect of amino density on the attachment, migration, and differentiation of rat neural stem cells In Vitro

Artificial extracellular matrices play important roles in the regulation of stem cell behavior. To generate materials for tissue engineering, active functional groups, such as amino, carboxyl, and hydroxyl, are often introduced to change the properties of the biomaterial surface. In this study, we chemically modified coverslips to create surfaces with different amino densities and investigated the adhesion, migration, and differentiation of neural stem cells (NSCs) under serum-free culture conditions. We observed that a higher amino density significantly promoted NSCs attachment, enhanced neuronal differentiation and promoted excitatory synapse formation in vitro. These results indicate that the amino density significantly affected the biological behavior of NSCs. Thus, the density and impact of functional groups in extracellular matrices should be considered in the research and development of materials for tissue engineering.     

Multiple density dependence in two sub-populations of the amphipod Monoporeia affinis: a potential for alternative equilibria

Possible mechanisms for differences in population densities and dynamics were investigated in the amphipod Monoporeia affinis at two deep sites in the northern Bothnian Sea. The two sites were sampled yearly for 10 years. Average sizes, growth and mortality of the different age-classes were estimated from the cohort structure of the two populations. Laboratory experiments also investigated the ability of the common predatory isopod Saduria entomon to cause densitydependent (DD) mortality of the prey M. affinis. At site A, 43 m depth, the average density of M. affinis was twice as high as at site B, 81 m depth. The fluctuations in density were asynchronous between the two sites. Recruitment and subadult sizes of Monoporeia affinis were density dependent at both sites. The main functional difference between the two populations seemed to be the DD mortality of the 1 + cohort that occurred only at the low-density site B. A corresponding DD mortality was found in the predation experiments at densities of 1 + m. affinis corresponding to those found at site B. The potential importance of the predator was also indicated by a significant negative correlation between the biomass of S. entomon and the rate of change in M. affinis density in the field. The similarities in the abiotic factors between the two sites suggested that differences in carrying capacity should be small. The results could be explained by the predation regulation hypothesis for the low-density population at site B, while at site A M. affinis seemed to be regulated by intra-specific competition and limited by predation. It is suggested that in this simple predator-prey system there is potential for the existence of alternative equilibria.     

MiR-216a-3p regulates the proliferation,apoptosis, migration,and invasion of lung cancer cells via targeting COPB2

ABSTRACT

Effect of miR-216a-3p on lung cancer hasn’t been investigated. Here, we explored its effects on lung cancer. MiR-216a-3p expression in lung cancer tissues and cells was detected by RT-qPCR. The target gene of miR-216a-3p was predicted by bioinformatics and confirmed by luciferase-reporter assay. After transfection, cell viability, migration, invasion, proliferation, and apoptosis were detected by MTT, scratch, transwell, colony formation, and flow cytometry. The expressions of COPB2 and apoptosis-related factors were detected by RT-qPCR or western blot. MiR-216a-3p was low-expressed and COPB2 was high-expressed in lung cancer tissues and cells. MiR-216a-3p targeted COPB2 and regulated its expression. MiR-216a-3p inhibited lung cancer cell viability, migration, invasion, and proliferation, while promoted apoptosis. Effect of miR-216a-3p on lung cancer was reversed by COPB2. MiR-216a-3p regulated proliferation, apoptosis, migration, and invasion of lung cancer cells via targeting COPB2.     

Met degradation by SAIT301, a Met monoclonal antibody,reduces the invasion and migration of nasopharyngeal cancer cells via inhibition of EGR-1 expression

Nasopharyngeal carcinoma (NPC) is a common malignant tumor with high invasive and metastatic potential. The hepatocyte growth factor (HGF)-Met signaling pathway has a critical role in mediating the invasive growth of many different types of cancer, including head and neck squamous cell carcinoma. HGF also stimulates NPC cell growth and invasion in the cell line model. In this study, we determined the inhibitory effect of Met, using a Met-targeting monoclonal antibody (SAIT301), on the invasive and growth potential of NPC cell lines. Met inhibition by SAIT301 resulted in highly significant inhibition of cell migration and invasion in both the HONE1 and HNE1 cell lines. In addition, we also found that co-treatment of SAIT301 and HGF decreased the anchorage-independent growth induced by HGF in HNE1 cell lines. After SAIT301 treatment, Met, together with its downstream signaling proteins, showed downregulation of p-Met and p-ERK, but not p-AKT, in both HONE1 and HNE1 cell lines. Interestingly, we found that HGF treatment of NPC cell lines induced early growth response protein (EGR-1) expression, which is involved in cell migration and invasion. In addition, co-treatment with SAIT301 and HGF inhibited the HGF-induced expression of EGR-1. Next, knockdown of EGR-1 using small-interfering RNA inhibited HGF-induced cell invasion in NPC cell lines, suggesting that the expression level of EGR-1 is important in HGF-induced cell invasion of NPC cells. Therefore, the results support that SAIT301 inhibited Met activation as well as the downstream EGR-1 expression and could have therapeutic potential in NPC. Taken together, we suggest that Met is an anticancer therapeutic target for NPC that warrants further investigation and clinical trials and SAIT301 may be a promising tool for NPC therapy.     

Comparative estimation of effective population sizes and temporal gene flow in two contrasting population systems

Estimation of effective population sizes (N(e)) and temporal gene flow (N(e)m, m) has many implications for understanding population structure in evolutionary and conservation biology. However, comparative studies that gauge the relative performance of N(e), N(e)m or m methods are few. Using temporal genetic data from two salmonid fish population systems with disparate population structure, we (i) evaluated the congruence in estimates and precision of long- and short-term N(e), N(e)m and m from six methods; (ii) explored the effects of metapopulation structure on N(e) estimation in one system with spatiotemporally linked subpopulations, using three approaches; and (iii) determined to what degree interpopulation gene flow was asymmetric over time. We found that long-term N(e) estimates exceeded short-term N(e) within populations by 2-10 times; the two were correlated in the system with temporally stable structure (Atlantic salmon, Salmo salar) but not in the highly dynamic system (brown trout, Salmo trutta). Four temporal methods yielded short-term N(e) estimates within populations that were strongly correlated, and these were higher but more variable within salmon populations than within trout populations. In trout populations, however, these short-term N(e) estimates were always lower when assuming gene flow than when assuming no gene flow. Linkage disequilibrium data generally yielded short-term N(e) estimates of the same magnitude as temporal methods in both systems, but the two were uncorrelated. Correlations between long- and short-term geneflow estimates were inconsistent between methods, and their relative size varied up to eightfold within systems. While asymmetries in gene flow were common in both systems (58-63% of population-pair comparisons), they were only temporally stable in direction within certain salmon population pairs, suggesting that gene flow between particular populations is often intermittent and/or variable. Exploratory metapopulation N(e) analyses in trout demonstrated both the importance of spatial scale in estimating N(e) and the role of gene flow in maintaining genetic variability within subpopulations. Collectively, our results illustrate the utility of comparatively applying N(e), N(e)m and m to (i) tease apart processes implicated in population structure, (ii) assess the degree of continuity in patterns of connectivity between population pairs and (iii) gauge the relative performance of different approaches, such as the influence of population subdivision and gene flow on N(e) estimation. They further reiterate the importance of temporal sampling replication in population genetics, the value of interpreting N(e)or m in light of species biology, and the need to address long-standing assumptions of current N(e), N(e)m or m models more explicitly in future research.     

Structural observations of time dependent oscillatory behavior of CuIICl2 solutions measured via extended X-ray absorption fine structure

Cell surface ECTO-NOX proteins exhibit a clock-related, temperature-independent entrainable pattern of periodic (24 min) oscillations in the rate of oxidation of NAD(P)H. Aqueous solutions of copper salts also oxidize NAD(P)H with a similar temperature-independent pattern. For both, five maxima are observed, two of which are separated by 6 min and the remaining three are separated by 4.5 min. In D2O, the pattern is retained but the period length is proportionately increased to 30 min in direct relationship to the 30 h circadian day observed with D2O-grown organisms. With copper solutions, periodic changes in redox potential correlate precisely with the periodic changes in the rates of NAD(P)H oxidation. Consequently, the local environment of the Cu2+ ion in copper chloride solutions was investigated by X-ray absorption spectroscopy. Detailed extended X-ray absorption fine structure (EXAFS) analyses revealed a pattern of oscillations closely resembling those of the copper-catalyzed oxidation of NADH. With CuCl2 in D2O, a pattern with a period length of 30 min was observed. The findings suggest a regular pattern of distortion in the axial and/or equatorial oxygen atoms of the coordinated water molecules which correlate with redox potential changes sufficient to oxidize NADH. A metastable equilibrium condition in the ratio of ortho to para nuclear spin orientation of the water associated hydrogen atoms would be kinetically consistent with a 24-30 min timeframe. The temperature independence of the biological clock can thus be understood as the consequence of a physical rather than a chemical basis for the timing events.     

The contact zone between two migration routes of silver fir,Abies alba (Pinaceae), revealed by allozyme studies

Enzyme data from 35 populations or regional samples ofAbies alba were obtained from different regions of Austria and Bavaria. The genetic analysis revealed a clear differentiation between the western and the north/eastern populations. A rather narrow transition zone (the river valleys of Salzach and Traun) with genetically intermediate populations connects these two regions. This west-east differentiation can be interpreted as the result of distinct migration routes from a glacial refuge in Central Italy. The transition zone is assumed to be the contact area of two routes north of the Alps. Each migration route of silver fir parallels the distribution range ofCardamine trifolia (east) andAposeris foetida (west) respectively. It is supposed that these two species were already associated with silver fir during colonization.Dedicated to emer. Univ.-Prof. DrFriedrich Ehrendorfer on the occasion of his 70th birthday     

Lymphocyte capping and lymphocyte migration as associated events in the in vivo antigen trapping process. an electron-microscopic autoradiographic study in the spleen of mice

The fate of 125I labeled antigen-antibody complexes in the first two hours after intravenous injection was followed in the spleen of mice by light and electron microscopic autoradiography. It appeared that AAC were phagocytized by granulocytes and by macrophages in the marginal zone and were present over the surface of lymphocytes in that area. By rinsing the spleen before fixation it could be shown that AAC were indeed bound to the lymphocyte membrane and not merely present in the blood plasma between the cells. The label was present in a spotty fashion or over a so-called uropod. The majority of labeled uropods (13 out of 17) pointed away from the follicle. From this it was inferred that these lymphocytes moved to and most probably into, the follicles. Inside the follicles, at 1 hour post injectionem, most of the label was associated with interfaces between lymphocytes. At 2 hours post injectionem there was a preferential localization over interfaces between lymphocytes and dendritic reticulum cells. It is conceivable that antigen that is introduced into the circulation is ultimately presented to dendritic reticulum cells in a complexed form with antibody, probably with complement, and with the B-cell receptor, since receptor shedding is a normal event following capping.     

Integrin-dependent neuroblastoma cell adhesion and migration on laminin is regulated by expression levels of two enzymes in the O-mannosyl-linked glycosylation pathway, PomGnT1 and GnT-Vb

O-mannosyl-linked glycans constitute a third of all brain O-linked glycoproteins, and yet very little is understood about their functions. Several congenital muscular dystrophies with central nervous system defects are caused by genetic disruptions in glycosyltransferases responsible for the synthesis of O-mannosyl glycans. The glycosyltransferase GnT-Vb, also known as GnT-IX, is expressed abundantly in the brain and testis and is proposed to be the enzyme that branches O-mannosyl-linked glycans. In this study, we show in a human neuronal model that GnT-Vb expression enhances neurite outgrowth on laminin. GnT-Vb has been shown to perform both N-linked and O-mannosyl-linked glycosylation. To determine if the effect on neurite outgrowth was due to N-linked or O-mannosyl-linked glycosylation by GnT-Vb we suppressed the expression of glycosyltransferases important for the elongation of both N-linked and O-mannosyl-linked glycans using RNA interference. Our results suggest that GnT-Vb and PomGnT1, enzymes involved in the O-mannosyl glycosylation pathway, play an active role in modulating integrin and laminin-dependent adhesion and migration of human neuronal cells.     

Temperature dependent development and survival of two sympatric species, Galerucella calmariensis and G. pusilla, on purple loosestrife

Two sympatric Europeanbeetles, Galerucella calmariensis (L.)and G. pusilla (Duft.) (Coleoptera:Chrysomelidae), have been released in NorthAmerica for biological control of purpleloosestrife, Lythrum salicaria L.(Lythraceae). Because establishment isaffected by environmental conditions, studieswere conducted at temperatures ranging from12.5 to 30 °C to determine differencesin rate of development and survival between thetwo species. No egg hatch occurred at12.5 °C for both species. Eggdevelopment was faster for G.calmariensis than for G. pusilla attemperatures 15 °C. Theminimum threshold temperature for eggdevelopment was lower for G. calmariensisthan G. pusilla. At 12.5 °C, G. calmariensis larvae developed 1.6 daysfaster than G. pusilla, but at 15, 20,25, and 27.5 °C G. pusilla developed2.9, 1.2, 1.1, and 0.6 days faster,respectively, than G. calmariensis. Nodifferences in developmental rate of pupa andtotal development time from egg to adulteclosion were observed for the two species. However, survival of G. calmariensisgenerally was higher than that of G.pusilla at lower temperatures. No differencewas observed in the preoviposition periodbetween the two species except at27.5 °C. The preoviposition period forboth species at 12.5 and 15 °C exceeded50 days and was much higher than for20 °C and above. The long preovipositionperiod suggests that temperatures below15 °C induce reproductive diapause. At15 °C, G. pusilla females livedlonger and also had a longer oviposition periodthan G. calmariensis. However, fasteregg and larval development, and higher survivalat 12.5 °C may give G. calmariensisa competitive advantage over G. pusillain cooler climates.     

Phytohormone influence on the expression of two isozyme systems in Nicotiana suaveolens calluses

Two types of calluses were obtained from seedlings of Nicotiana suaveolens when cultured on three different media, two of them supplemented with benzyl-adenine (BA) and naphthalen acetic acid (type B calluse), and one with only naphthalen acetic acid (type N calluses). Two isozyme systems were studied in these calluses which showed differences in their regenerative capacity, shoot formation being mediated by benzyl-adenine. Differences in the peroxidases (PX) were not found, but there were in the case of glutamic-oxaloacetic-transaminases (GOT), the isozyme patterns of which presented differences between both types of calluses. These changes observed could be a reflex of different developmental stages in both types of calluses.     

Distinct contributions of CXCR4b and CXCR7/RDC1 receptor systems in regulation of PGC migration revealed by medaka mutants kazura and yanagi

Migratory pathways of PGCs to the gonad vary depending on the vertebrate species, yet the underlying regulatory mechanisms guiding PGCs are believed to be largely common. In teleost medaka embryo, PGC migration follows two major steps before colonizing in gonadal areas: (1) bilateral lineup in the trunk and (2) posterior drift of PGCs. kazura (kaz) and yanagi (yan) mutants of medaka isolated in mutagenesis screening were defective in the first and second steps, respectively. kaz^j2-15D was identified as a missense mutation in chemokine receptor gene cxcr4b expressed in PGCs. Embryonic injection of cxcr4b mRNA with vasa 3′ UTR rescued the PGC phenotype of kaz mutant, indicating a cell-autonomous function of cxcr4b in PGCs. yan^j6-29C was identified as a nonsense mutation in the cxcr7/rdc1 gene encoding another chemokine receptor. cxcr7 transgene with genomic flanking sequences rescued the yan mutant phenotype efficiently at the G0 generation. cxcr7 was expressed in somites rather than PGCs. cxcr7-expressing somitic domain expanded posteriorly with its margin immediately anterior of posteriorly drifting PGCs, as if PGCs were thrusted toward the gonadal area. kaz and yan mutants are also defective in lateral line positioning, suggesting combined employment of these receptor systems in various cell migratory processes.     

Enhanced suppression of proliferation and migration in highly-metastatic lung cancer cells by combination of valproic acid and coumarin-3-carboxylic acid and its molecular mechanisms of action

Valproic acid (VPA) as a broad-spectrum inhibitor of histone deacetylase, has been used in cancer therapy. Recently, the combination of VPA with other anticancer agents has been considered as a useful and necessary strategy to inhibit tumor growth and progression. The coumarin derivates from natural plants have been shown to be the promising natural anticancer agents. However, no literature is available on the anticancer effects of the combination of VPA and coumarin-3-carboxylic acid (HCCA). Here we show that this combination significantly increases inhibitory effects against the proliferation and migration in highly-metastatic lung cancer cells by inducing apoptosis and cell cycle arrest as well as regulating related protein expressions. Our results indicate that this combination of VPA with HCCA not only enhances the protein levels of Bax, cytosolic cytochrome c, caspase-3 and PARP-1 but also reduces the protein expressions of Bcl-2, cyclin D1 and NF-κB as well as inhibits the phosphorylation and expressions of Akt, EGFR, VEGFR2 and c-Met in the cancer cells. Our results suggest that the combination of VPA with HCCA suppresses the proliferation and migration of lung cancer cells via EGFR/VEGFR2/c-Met-Akt-NF-κB signaling pathways; this combination may have a wide therapeutic and/or adjuvant therapeutic application in the treatment of lung cancer.     

Intensive measurement of nitrous oxide emissions from a corn–soybean–wheat rotation under two contrasting management systems over 5 years

No‐tillage (NT), a practice that has been shown to increase carbon sequestration in soils, has resulted in contradictory effects on nitrous oxide (N₂O) emissions. Moreover, it is not clear how mitigation practices for N₂O emission reduction, such as applying nitrogen (N) fertilizer according to soil N reserves and matching the time of application to crop uptake, interact with NT practices. N₂O fluxes from two management systems [conventional (CP), and best management practices: NT + reduced fertilizer (BMP)] applied to a corn (Zea mays L.), soybean (Glycine max L.), winter‐wheat (Triticum aestivum L.) rotation in Ontario, Canada, were measured from January 2000 to April 2005, using a micrometeorological method. The superimposition of interannual variability of weather and management resulted in mean monthly N₂O fluxes ranging from − 1.9 to 61.3 g N ha⁻¹ day⁻¹. Mean annual N₂O emissions over the 5‐year period decreased significantly by 0.79 from 2.19 kg N ha⁻¹ for CP to 1.41 kg N ha⁻¹ for BMP. Growing season (May–October) N₂O emissions were reduced on average by 0.16 kg N ha⁻¹ (20% of total reduction), and this decrease only occurred in the corn year of the rotation. Nongrowing season (November–April) emissions, comprised between 30% and 90% of the annual emissions, mostly due to increased N₂O fluxes during soil thawing. These emissions were well correlated (r²= 0.90) to the accumulated degree‐hours below 0 °C at 5 cm depth, a measure of duration and intensity of soil freezing. Soil management in BMP (NT) significantly reduced N₂O emissions during thaw (80% of total reduction) by reducing soil freezing due to the insulating effects of the larger snow cover plus corn and wheat residue during winter. In conclusion, significant reductions in net greenhouse gas emissions can be obtained when NT is combined with a strategy that matches N application rate and timing to crop needs.